The aim of thesis is focused on isolation of DNA in quality for polymerase chain reaction (PCR) and the identification of probiotic bacteria. From six probiotic supplements for children were isolated PCR-ready DNAs using magnetic carriers P(HEMA-co-GMA). Isolated DNA was amplified by genus-specific and species-specific primers. DNAs of Lactobacillus, Bifidobacterium and Streptococcus genera were identified as: L. acidophilus, L. rhamnosus, L. casei, B. bifidum, B. longum ssp. longum, B. breve, B. longum ssp infantis, B. animalis and S. thermophilus. The identification corresponded with the data declared by the producers.

Made available in DSpace on 2015-04-20T12:31:28Z (GMT). No. of bitstreams: 1 Isabel da Mota Pontes.pdf: 2179836 bytes, checksum: 0b2f33d44ee029fbf560b3505ea884b2 (MD5) Previous issue date: 2009-12-14 / The present work presents the development of a new system called multiplex Pentaplex-ised, composed of 5 microsatellite loci STR type (D5S198498, D3S18773, GH15, D8S82535 and D11S118590). Such microsatellites have been identified in the human genome, using the program BLAST-N (GenBank). Specific primers were designed for amplification via PCR. Fluorescent primers were synthesized to allow simultaneous analysis of multiple amplicons (multiplex analysis). All microsatellite loci were genotyped confirming the high degree of polymorphism. In the validation, the minimum DNA concentration of for amplification was estimated at 0.25 ng. Several types of biological samples (blood, saliva, urine and semen) were acquired satisfactorily. We built a database of microsatellite allele frequencies for three Brazilian populations -Amazonas, Rio de Janeiro and Espírito Santo. The distributions of allele frequencies showed little variation between population groups. By means of statistical tests of χ2 using the program Arlequim 3.1 most alleles do not adhere to Hardy-Weinberg equilibrium if performed separately for each of the populations. However, in the analysis performed with the whole set of three populations, there is high adherence to the Hardy-Weinberg equilibrium. The three populations were grouped and compared pair-to-pair, showing that the genetic variation is greater within populations than between them. Statistical data about the power of exclusion, discrimination power and polymorphism information content for the five loci were determined for each STR and together to assess the potential of the system Pentaplex-ised in human identification. We observed that this multiplex system is suitable for human identification and that can be used in a complementary method with other systems for genetic analysis, by having a cumulative power of discrimination of 99.996% and 93.436% cumulative exclusion / O presente trabalho consiste no desenvolvimento de um novo sistema multiplex denominado PentaPlex-ISED, composto por 5 loci tipo microssatélites STR (D5S198498, D3S18773, GH15, D8S82535 e D11S118590). Esses microssatélites foram identificados no genoma humano, por meio do programa BLAST-N (GenBank). Primers específicos foram desenhados para amplificação via PCR. Primers fluorescentes foram sintetizados para permitir análise simultânea de vários amplicons (análise multiplex). Todos os loci microssatélites foram genotipados confirmando-se o alto grau de polimorfismo. Na validação do sistema, a concentração mínima de DNA para a amplificação dos loci incluídos nesse multiplex foi estimada em 0,25 ng. Diversos tipos de amostras biológicas (sangue, saliva, urina e sêmen) foram adquiridas satisfatoriamente. Foi construído um banco de dados de freqüências dos alelos microssatélites para três populações brasileiras (Amazonas, Rio de Janeiro e Espírito Santo). Confirmou-se a independência dos loci. As distribuições das freqüências alélicas apresentaram pouca variação entre os grupos populacionais. Por meio dos testes estatísticos de χ2, utilizando o programa Arlequim 3.1 verificou-se que a maioria dos loci não adere as condições do equilíbrio de Hardy-Weinberg se análise for feita separadamente em cada uma das populações. No entanto, na análise feita com o conjunto total das três populações, há aderência ao Equilíbrio de Hardy-Weinberg. As três populações foram agrupadas e comparadas par-a-par; mostrando-se que a variação genética é maior dentro das populações que entre elas. Os dados estatísticos sobre o poder de exclusão, poder de discriminação e o conteúdo de informação do polimorfismo, para os 5 loci foram determinados para cada STR e em conjunto para avaliar o potencial do sistema PentaPlex-ISED na identificação humana. Observou-se que este sistema multiplex é adequado para identificação humana e que pode ser usado de forma complementar com os outros sistemas de análises genéticas, por possuir um poder de discriminação acumulado em 99,996 % e o poder de exclusão acumulado em 93,436 %.

La criminalistique peut être définie comme l’application de procédés techniques aux investigations judiciaires permettant l’étude scientifique des traces et des indices retrouvés sur les scènes de crime.Depuis la découverte de l’empreinte génétique par Sir Alec Jeffreys en 1984, le monde judiciaire s’est profondément ancré dans l’ère de l’ADN en raison d’évolutions technologiques successives dans le domaine de la biologie moléculaire et ses applications en criminalistique. Le besoin de réponse instantanée est omniprésent dans les esprits. La mise en œuvre de techniques d’analyses simples, sensibles, fiables et permettant d’obtenir des résultats dans les plus brefs délais sont les clés du succès.Au cours des processus techniques, la collecte du matériel biologique, et donc de l’ADN au sein de la trace, constitue une étape incontournable et cruciale qui va conditionner la réussite des analyses. Ce travail de recherche a donc consisté à développer des solutions performantes de prélèvements de matériels biologiques soit en détournant de leur fonction initiale des solutions existantes soit en développant des solutions simples mais innovantes combinant les avantages des solutions existantes. Ces travaux ont permis de donner naissance au micro-écouvillon GendSAG. Les potentialités de GendSAG permettent de proposer une solution alternative aux solutions commerciales de systèmes intégrés d’analyses rapide d’ADN. Cette solution alternative d’analyse rapide et haut débit de l’ADN mise en œuvre dans un laboratoire mobile au plus près de la scène de crime répond non seulement à la grande majorité des avantages des systèmes intégrés mais également à toutes leurs limitations. / Forensic sciences can be defined as the used of technical processes to judicial investigations allowing the scientific study of traces and evidences found on crime scenes.Since the discovery of DNA fingerprinting by Sir Alec Jeffreys in 1984, the legal world has become deeply rooted in the DNA by successive technological developments in molecular biology and its applications in forensic. The need for instant response is omnipresent in the minds. The key to success is the implementation of simple, sensitive, reliable analytical techniques that enable results to be achieved in the shortest possible time.During these technical processes, the collection of biological samples, is an unavoidable and a crucial step that will condition the analysis success rate. This study consisted in developing efficient biological collecting solutions either by diverting from their original function the existing solutions or by developing simple but innovative solutions combining the advantages of the existing solutions. This allowed developing the micro-swab GendSAG. The potentialities of GendSAG make it possible to propose an alternative solution to the commercial rapid DNA analysis integrated systems. This rapid, cost effective and high-throughput DNA analysis solution performed in a dedicated mobile laboratory directly into the crime scene enables the large majority of the rapid DNA analysis integrated systems benefits and also all of their limitations.